Natural killer (NK) cells are operationally defined as cells capable of mediating spontaneous in vitro cytotoxicity against a variety of target cell populations without prior sensitization or MHC restriction. They express NKH1 while classic CTL, B cells, and myeloid cells do not. Their in vivo function, lineage, and site of maturation remain unknown. NK cells express CD2, and each can undergo activation through the CD2 pathway. The majority of peripheral blood CD2-CD3-NKH1-cells lack TCR gene rearrangement suggesting either a non-T cell lineage or a common precursor cell which is prethymic. NK cells are the first lymphocytes to return following T cell depleted bone marrow transplant, but the significance remains unclear. Freshly isolated human thymocytes lack expression of NKH1 and non-MHC- restricted cytotoxicity, but short term culture in rIL-2 results in the induction of NKH1 expression on NKH1- thymocytes and the development of NK killing. CD3-NKH1+ thymocytes can be generated from such cultures, but the precursor population is unknown. A subpopulation of early human thymocytes (CD2+CD3-) express IL-2R, produce IL-2 autocrine pathway, which may lead to proliferation and differentiation of functionally diverse cells. This early thymocyte may lead to proliferation and differentiation of functionally diverse cells. This early thymocyte may serve as a precursor for the IL-2 cultured CD3-NKH1+ NK cell and possibly for the CD3-NKH1+ NK cell found in human peripheral blood. Several possibilities may exist: 1) Within the thymus, the CD2+CD3- thymocyte is committed to expression of the TCR gene products. With continued exposure to IL-2 in vivo, some progeny differentiate into CD3+NKH1+ NK cells only, but no CD3-NKH1+ cells are thymic-derived in vivo 2) The CD2+CD3- thymocyte is not yet committed to express the TCR gene products, and with continued exposure to IL-2 in vivo, a fraction of thymocyte progeny differentiate into CD3\NKH1+ NK cells within human thymus and exit to the periphery. 3) A population of CD2+CD3- progenetor cells exist outside the thymus and its therefore not able to rearrange TCR genes. Through an IL-2 autocrine pathway, functional development of the CD3-NKH1+ cell would occur without TCR gene rearrangement or expression. Specific aims in phase I of this proposal involve an extensive in vitro phenotypic functional and molecular characterization of early thymocyte subpopulations, their progeny, and the CD3-NKH1+ cultured thymocytes. The data obtained should offer significant insights into the possible origin(s) and lineage(s) of human NK cells. Phase II will focus on further characterization of the NKH1 antigen and growth factors.